IGF::OT::IGF MULTIPLEXED HIGH-CONTENT ASSAY TO MEASURE CELLULAR STRESS MECHANISMS FOR PREDICTIVE TOXICOLOGY STUDIES

承包商目前提供了一个高通量分析平台，通过其独特的能力，实时监测活细胞中以前无法获得的生物学的多个方面，快速确定化合物的作用机制和毒性责任。 通过这项SBIR第二阶段合同，承包商现有的平台将被应用于开发一种新型的活细胞多重荧光检测，该检测报告每孔至少10个不同的细胞应激和细胞器稳态读数。将生成一组细胞应激报告基因，并将其稳定整合到与化合物毒性评价相关的细胞系中。所得到的一组稳定的报告细胞系将用作我们的多重测定的基础。我们的测定将通过测试其对已知诱导细胞应激的工具化合物的响应来验证。我们的检测方法对高通量筛选的适用性将通过运行10，000个样本的中试筛选进行评估，然后在384孔板格式中扩展到至少100，000个样本/天的通量，并实现完全的检测自动化。 承包商提出的检测方法将通过减少确定药物和环境化学品毒性的时间和成本来有益于公众健康，从而提高我们识别有益治疗方法的能力，并积极主动地管理有害的环境毒素。

The Contractor currently provides a high-throughput assay platform that rapidly pinpoints compound mechanism of action and toxicity liabilities through its unique ability to monitor multiple facets of previously inaccessible biology in live cells in real-time. Through this SBIR Phase II contract, the Contractor's existing platform will be applied to the development of a novel live-cell multiplexed fluorescent assay that reports an aggregate of at least 10 distinct readouts of cellular stress and organelle homeostasis per well. A panel of multicolor cell stress reporters will be generated and stably integrated into cell lines relevant for compound toxicity evaluation. The resulting panel of multicolor stable reporter cell lines will be used as the foundation for our multiplexed assay. Our assay will be validated by testing its response to tool compounds known to induce cellular stress. Our assay's suitability for high-throughput screening will be evaluated by running a 10,000-sample pilot screen, followed by scaling to at least 100,000 samples/day throughput in 384-well plate format with full assay automation. The Contractor's proposed assay will benefit public health by reducing the time and cost to pinpoint toxicities of pharmaceutical and environmental chemicals, thus improving our ability to identify beneficial therapeutics, and to proactively manage harmful environmental toxins.